Lung cancer is the leading cause of death from cancer and kills over 150,0000 people each year in the United States. Since early stage lung cancer can be cured by surgical resection, efforts are made to develop effective screening approaches for early stage disease. DNA methylation biomarkers hold great promise as molecular tools to identify early stage lung tumors. Only a limited panel of genes has been identified that are methylated at a high frequency in early stage lung cancers. We have begun to develop new technology applicable for genome-wide analysis of DNA methylation changes in cancer. The methylated fraction of the genome is selectively enriched using the methylated-CpG island recovery assay (MIRA), which is based on the high affinity of the MBD2/MBD3L1 complex for methylated DNA. This approach can be combined with microarrays to identify genome-wide methylation changes in cancer. In this application, we will further develop and apply the MIRA approach for microarray-based screening of genome-wide DNA methylation differences between normal lung tissue and early stage lung tumors. The reason for focusing on early stages is that any identified new methylation biomarker may be appropriate for early detection and diagnosis. DNA isolated from different histological types of lung tumors will be used for hybridization on different CpG island and promoter microarray platforms. Genes identified as being methylated in early stage lung tumors by microarray analysis, will be verified in a larger set of samples using standard methylation detection techniques. The technology will be further developed to be applicable to small numbers of cells (<1000) and to tumor biospecimens including frozen, embedded, and formalin-fixed, paraffin embedded tissues. DNA methylation biomarkers hold great promise as molecular tools to identify early stage lung tumors. However, only a limited number of genes have been identified that are methylated in early stage lung cancers. We have been developing a new technology applicable to genome- wide screening of DNA methylation changes in cancer. The methylated fraction of the genome is selectively enriched using the Methylated-CpG Island Recovery Assay (MIRA). MIRA has shown initial promise to be a specific and general methylation detection tool and is compatible with microarray analysis. In this R21 application, we will further refine the methodology and attempt to apply the MIRA approach for microarray-based screening of genome-wide DNA methylation differences between normal lung tissue and early stage lung tumors. [unreadable] [unreadable] [unreadable]